Cargando…

Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners

To characterize our recently established in vitro glaucomatous human trabecular meshwork (HTM) models using dexamethasone (DEX)- or TGF-β2-treated HTM cells, (1) two-dimensional (2D) cultured HTM cells were characterized by means of the real-time cellular metabolism analysis using a Seahorse analyze...

Descripción completa

Detalles Bibliográficos
Autores principales: Watanabe, Megumi, Sato, Tatsuya, Tsugeno, Yuri, Umetsu, Araya, Suzuki, Soma, Furuhashi, Masato, Ida, Yosuke, Hikage, Fumihito, Ohguro, Hiroshi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9219943/
https://www.ncbi.nlm.nih.gov/pubmed/35740359
http://dx.doi.org/10.3390/biomedicines10061338
_version_ 1784732247701585920
author Watanabe, Megumi
Sato, Tatsuya
Tsugeno, Yuri
Umetsu, Araya
Suzuki, Soma
Furuhashi, Masato
Ida, Yosuke
Hikage, Fumihito
Ohguro, Hiroshi
author_facet Watanabe, Megumi
Sato, Tatsuya
Tsugeno, Yuri
Umetsu, Araya
Suzuki, Soma
Furuhashi, Masato
Ida, Yosuke
Hikage, Fumihito
Ohguro, Hiroshi
author_sort Watanabe, Megumi
collection PubMed
description To characterize our recently established in vitro glaucomatous human trabecular meshwork (HTM) models using dexamethasone (DEX)- or TGF-β2-treated HTM cells, (1) two-dimensional (2D) cultured HTM cells were characterized by means of the real-time cellular metabolism analysis using a Seahorse analyzer, and (2) the effects of mechanical compression stresses toward the three-dimensional (3D) HTM spheroids were evaluated by analyzing the gene expression of several ECM proteins, inflammatory cytokines, and ER stress-related factors of those 3D HTM spheroid models. The results indicated that (1) the real-time cellular metabolism analysis indicated that TGF-β2 significantly induced an energy shift from mitochondrial oxidative phosphorylation (OXPHOS) into glycolysis, and DEX induced similar but lesser effects. In contrast, ROCK2 inhibition by KD025 caused a substantial reverse energy shift from glycolysis into OXPHOS. (2) Upon direct compression stresses toward the untreated control 3D HTM spheroids, a bimodal fluctuation of the mRNA expressions of ECM proteins was observed for 60 min, that is, initial significant upregulation (0–10 min) and subsequent downregulation (10–30 min) followed by another upregulation (30–60 min); those of inflammatory cytokines and ER stress-related factors were also bimodally changed. However, such compression stresses for 30 min toward TGF-β2- or DEX-treated 3D HTM spheroids induced downregulation of most of those of inflammatory cytokines and ER stress-related factors in addition to upregulation of COL1 and downregulation of FN. The findings presented herein indicate that (1) OXPHOS of the HTM cells was decreased or increased by TGF-β2 or DEX stimulation or ROCK2 inhibition, and (2) mechanical compression stresses toward 3D HTM spheroids may replicate acute, subacute, and chronic HTM models affected by elevated intraocular pressures.
format Online
Article
Text
id pubmed-9219943
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-92199432022-06-24 Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners Watanabe, Megumi Sato, Tatsuya Tsugeno, Yuri Umetsu, Araya Suzuki, Soma Furuhashi, Masato Ida, Yosuke Hikage, Fumihito Ohguro, Hiroshi Biomedicines Article To characterize our recently established in vitro glaucomatous human trabecular meshwork (HTM) models using dexamethasone (DEX)- or TGF-β2-treated HTM cells, (1) two-dimensional (2D) cultured HTM cells were characterized by means of the real-time cellular metabolism analysis using a Seahorse analyzer, and (2) the effects of mechanical compression stresses toward the three-dimensional (3D) HTM spheroids were evaluated by analyzing the gene expression of several ECM proteins, inflammatory cytokines, and ER stress-related factors of those 3D HTM spheroid models. The results indicated that (1) the real-time cellular metabolism analysis indicated that TGF-β2 significantly induced an energy shift from mitochondrial oxidative phosphorylation (OXPHOS) into glycolysis, and DEX induced similar but lesser effects. In contrast, ROCK2 inhibition by KD025 caused a substantial reverse energy shift from glycolysis into OXPHOS. (2) Upon direct compression stresses toward the untreated control 3D HTM spheroids, a bimodal fluctuation of the mRNA expressions of ECM proteins was observed for 60 min, that is, initial significant upregulation (0–10 min) and subsequent downregulation (10–30 min) followed by another upregulation (30–60 min); those of inflammatory cytokines and ER stress-related factors were also bimodally changed. However, such compression stresses for 30 min toward TGF-β2- or DEX-treated 3D HTM spheroids induced downregulation of most of those of inflammatory cytokines and ER stress-related factors in addition to upregulation of COL1 and downregulation of FN. The findings presented herein indicate that (1) OXPHOS of the HTM cells was decreased or increased by TGF-β2 or DEX stimulation or ROCK2 inhibition, and (2) mechanical compression stresses toward 3D HTM spheroids may replicate acute, subacute, and chronic HTM models affected by elevated intraocular pressures. MDPI 2022-06-06 /pmc/articles/PMC9219943/ /pubmed/35740359 http://dx.doi.org/10.3390/biomedicines10061338 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Watanabe, Megumi
Sato, Tatsuya
Tsugeno, Yuri
Umetsu, Araya
Suzuki, Soma
Furuhashi, Masato
Ida, Yosuke
Hikage, Fumihito
Ohguro, Hiroshi
Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title_full Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title_fullStr Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title_full_unstemmed Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title_short Human Trabecular Meshwork (HTM) Cells Treated with TGF-β2 or Dexamethasone Respond to Compression Stress in Different Manners
title_sort human trabecular meshwork (htm) cells treated with tgf-β2 or dexamethasone respond to compression stress in different manners
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9219943/
https://www.ncbi.nlm.nih.gov/pubmed/35740359
http://dx.doi.org/10.3390/biomedicines10061338
work_keys_str_mv AT watanabemegumi humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT satotatsuya humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT tsugenoyuri humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT umetsuaraya humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT suzukisoma humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT furuhashimasato humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT idayosuke humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT hikagefumihito humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners
AT ohgurohiroshi humantrabecularmeshworkhtmcellstreatedwithtgfb2ordexamethasonerespondtocompressionstressindifferentmanners